title: Detection of Papaya leaf distortion mosaic virus by reverse-transcription loop-mediated isothermal amplification.
abstract: Papaya leaf distortion mosaic virus (PLDMV) can infect transgenic papaya resistant to a related pathogen, Papaya ringspot virus (PRSV), posing a substantial threat to papaya production in China. Current detection methods, however, are unable to be used for rapid detection in the field. Here, a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of PLDMV, using a set of four RT-LAMP primers designed based on the conserved sequence of PLDMV CP. The RT-LAMP method detected specifically PLDMV and was highly sensitive, with a detection limit of 1.32×10(-6) μg of total RNA per reaction. Indeed, the reaction was 10 times more sensitive than one-step RT-PCR, while also requiring significantly less time and equipment. The effectiveness of RT-LAMP and one-step RT-PCR in detecting the virus were compared using 90 field samples of non-transgenic papaya and 90 field samples of commercialized PRSV-resistant transgenic papaya from Hainan Island. None of the non-transgenic papaya tested positive for PLDMV using either method. In contrast, 19 of the commercialized PRSV-resistant transgenic papaya samples tested positive by RT-LAMP assay, and 6 of those tested negative by RT-PCR. Therefore, the PLDMV-specific RT-LAMP is a simple, rapid, sensitive, and cost-effective tool in the field diagnosis and control of PLDMV.
authors.affiliation: Key Laboratory of Biology and Genetic Resources of Tropical Crops, Ministry of Agriculture, Institute of Tropical Bioscience and Biotechnology & Analysis and Testing Center, Chinese Academy of Tropical Agricultural Sciences, Haikou 571101, China.
authors.forename: Wentao
authors.initials: W
authors.lastname: Shen
authors.forename: Decai
authors.initials: D
authors.lastname: Tuo
authors.forename: Pu
authors.initials: P
authors.lastname: Yan
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Li
authors.forename: Peng
authors.initials: P
authors.lastname: Zhou
mesh_terms: D029441:Carica / Q000821:virology*; D002681:China; D003362:Cost-Benefit Analysis; D017931:DNA Primers / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000379:methods*; D010935:Plant Diseases / Q000821:virology*; D017800:Potyvirus / Q000235:genetics / Q000302:isolation & purification*; D048348:Reverse Transcription; D012680:Sensitivity and Specificity; D013997:Time Factors; D014773:Virology / Q000191:economics / Q000379:methods*
chemical_list: D017931:DNA Primers
keywords: PLDMV detection; Papaya leaf distortion mosaic virus; RT-PCR; Reverse-transcription loop-mediated isothermal amplification